This revised application describes a rigorous training program of education and investigation to facilitate the applicant's development into an independent physician-scientist in the field of anemia of chronic renal failure (CRF). The applicant, Dr. Saul Nurko, is a nephrologist at the Cleveland Clinic, with a principal clinical interest in CRF and its consequences. Training will be supervised by the Sponsor, Dr. Paul Fox. His laboratory in the Dept. of Cell Biology investigates cellular and molecular mechanisms regulating iron homeostasis, and defects that lead to anemia. Dr. Linda Graham, project Co-Sponsor, is a vascular surgeon who will have primary responsibility for training and performance in studies using the surgical model of CRF in the mouse. Anemia is a common and debilitating consequence of advanced kidney disease. The advent of recombinant human erythropoietin (Epo) treatment significantly changed how nephrologists manage anemia in this population, and has led to better outcomes and a higher quality of life for most patients. However, the response to Epo is variable and up to 10 percent of the dialysis population are "Epo-resistant". Variations in Epo-responsiveness may be due to physiological factors including uremia, inflammation, iron status, and environmental and genetic factors. We propose to investigate the expression of genes influencing the response to Epo during CRF. We have begun a microarray-based approach to identify candidate genes with altered expression during CRF induction or after treatment with Epo. A mouse model of surgery-induced CRF was selected which gives a state of uremia and anemia comparable to human disease. In Preliminary Studies, mice were subjected to CRF for 3 wk, and then half were treated with Epo for an additional 3 wk. Systemic responses were as described previously, namely, induction of severe uremia and anemia. Treatment with Epo partially restored hematological parameters in most mice. At sacrifice, RNA was isolated from liver, spleen, and bone marrow. Liver RNA from 2 mice in each group was analyzed using the Affymetrix GeneChip system. CRF induced the expression of several peroxisome proliferator-activated receptor (PPAR)-alpha-inducible genes including cytochrome p450 type A10 and insulin-like growth factor binding protein (IGFBP)-1. To our knowledge this is the first evidence for activation of hepatic PPAR-alpha by CRF. Our preliminary studies have led us to propose the following hypothesis: Activation of liver PPAR-alpha during CRF, possibly via uremic toxins in blood, alters the expression of specific hepatic genes. Dysregulation of these genes inhibits erythropoiesis, particularly, in response to Epo. We will test this hypothesis by pursuing the following Specific Aims: (1) Determine the effects of CRF on the expression of PPAR-(-induced and erythropoiesis-related genes in liver (2) determine the relationship between the extent of CRF and PPAR-alpha activity and erythropoiesis in mice with CRF, and (3) determine the in vivo role of PPAR-alpha in CRF-mediated alteration of gene expression using PPAR-alpha-null and clofibrate-treated mice. We expect that these studies will help us to understand the molecular mechanisms involved in Epo-resistance during CRF, and possibly will lead to the development of therapeutic agents that overcome this resistance and potentiate the activity of Epo.